This application for a Mentored Clinical Scientist Development Award (K08) for Dr. Ivona Percec, Assistant Professor in the Dept. of Surgery at the University of Pennsylvania, proposes a thorough investigation into the epigenetic factors that regulate primary human adipose aging. This proposal will fulfill the educational objective of the development award by facilitating the expansion of Dr. Percec's knowledge base into novel lines of inquiry through mentorship in these new areas. The mentors assisting in this grant proposal, Drs. Ken Zaret and Shelley Berger, are world experts in the epigenetic regulation of cellular differentiation and aging, respectively, and will provide essential guidance for the successful completion of both the educational mission of the award as well as the proposed research plan. Additional collaborators, Drs. Brad Johnson, Bob Pignolo and Patrick Seale will further complement and advance Dr. Percec's education in gerontology and stem cell biology. This K08 award will enable Dr. Percec to fulfill her long-term goal of becoming a leading independent investigator in the epigenetics of aging and the biology of human adipose tissue. The goal of this proposal is to investigate the function of the sirtuin genes in aging human primary adipose tissue, to identify critical sirtuin target genes and transcriptional networks specifically regulating aging of human adipose tissue, and to modulate sirtuin activity in primary adipose stem cells to improve age-related decline in cellular functions. While adipose tissue plays a central role in metabolism, caloric restriction pathways, and is the largest known reservoir of adult stem cells, the epigenetic factors resulting in normal human adipose aging are poorly understood. I hypothesize that the sirtuin genes contribute significantly to adipose tissue aging and found in my preliminary studies that SIRT1, SIRT3, SIRT6, and SIRT7 are differentially expressed in aging adipose tissue. My recent data suggest that specific sirtuin target proteins express predicted age-related changes in acetylation status. Based on these studies, I propose to validate the functional role of sirtuins in primary human adipose aging and to identify specific sirtuin regulators that reverse or induce age-associated phenotypes in human primary adipose-derived stem cells. Specifically, the proposed work will: 1) validate a functional role for sirtuins in human adipose aging by examining the acetylation status of sirtuin target proteins in young versus old human adipose tissue; 2) establish critical sirtuin chromatin occupancy in response to the onset of aging in human adipose aging by performing sirtuin ChIP-seq analyses in young versus old primary adipocytes, stromal vascular fractions and adipose-derived stem cells; 3) determine whether sirtuin dosage modulation in adipose stem cells can attenuate age-related decline in cellular functions. Understanding the mechanisms of sirtuin gene regulation of human aging through the study of primary human adipose tissue will shed light on the basis of human cellular and tissue aging and have profound implications for regenerative medicine applications.